Figure 2

p63 depletion attenuates ATM expression and ATM-dependent phosphorylation. (A) pSUPER-p63 RNAi attenuates ATM mRNA levels. HaCat cells were transfected with 1 μg pSUPER-CON or pSUPER-p63si vectors, and selected for geneticin resistance for 4 days. mRNA was extracted from surviving cells and real-time RT-PCR was used to quantitate changes in ATM mRNA expression. Data is represented as fold-change over empty vector control. (B) pSUPER-p63si inhibits ATM expression and p53 Serine-15 phosphorylation in HaCat cells. pSUPER-CON or pSUPER-p63si transfected HaCat cells were harvested after selection for 48 and 96 hours, and immunoblotted for the indicated proteins. (C) pSUPER-p63 RNAi inhibits HaCat cell proliferation. HaCaT cells were transfected with pSUPER-CON or pSUPER-p63si vectors, selected for geneticin resistance for 14 days, and surviving colonies were giemsa stained and counted. (D) siRNA-mediated p63 depletion attenuates ATM mRNA expression. HaCat cells untreated (NT) or treated with control siRNA or p63-targeted siRNA were harvested after 24 hrs or 48hrs and analysed by real-time RT-PCR for p63 and ATM mRNA expression. Data is normalised to β-actin and represented as fold-change over mRNA level in cells treated with control siRNA in particular time point. The data represent the average of three independent experiments + SD. The symbol asterisk denotes significant difference with p values < 0.05 determined by Mann-Whitney test, from the cells treated with control siRNA.